Compounds having biological activity can be identified by screening diverse collections of compounds (i.e. libraries of compounds) produced through synthetic chemical techniques. Such screening methods include methods wherein the library comprises a plurality of compounds synthesized at specific locations on the surface of a solid support whereby a receptor is appropriately labelled to bind to and identify a compound, e.g., fluorescent or radioactive labels. Correlation of the labelled receptor bound to the support and its location on the support identifies the binding compound (U.S. Pat. No. 5,143,854).
Central to these methods is the screening of a multiplicity of compounds in the library and the ability to identify the structures of the compounds which have a requisite biological activity. In order to facilitate synthesis and identification, the compounds in the library are typically formed on solid supports. Usually each such compound is covalently attached to the support via a cleavable or non-cleavable linking arm. The libraries of compounds can be screened either on the solid support or as cleaved products to identify compounds having good biological activity.
A particular class of compounds which would be useful for inclusion in screening libraries are pyrrolobenzodiazepines (PBDs). PBDs have the ability to recognise and bond to specific sequences of DNA; the most preferred sequence is PuGPu (Purine-Guanine-Purine). The first PBD antitumour antibiotic, anthramycin, was discovered in 1965 (Leimgruber et al., 1965 J. Am. Chem. Soc., 87, 5793-5795; Leimgruber et al., 1965 J. Am. Chem. Soc., 87, 5791-5793). Since then, a number of naturally occurring PBDs have been reported, and over 10 synthetic routes have been developed to a variety of analogues (Thurston et al., 1994 Chem. Rev. 1994, 433-465). Family members include abbeymycin (Hochlowski et al., 1987 J. Antibiotics, 40, 145-148), chicamycin (Konishi et al., 1984 J. Antibiotics, 37, 200-206), DC-81 (Japanese Patent 58-180 487; Thurston et al., 1990, Chem. Brit., 26, 767-772; Bose et al., 1992 Tetrahedron, 48, 751-758), mazethramycin (Kuminoto et al., 1980 J. Antibiotics, 33, 665-667), neothramycins A and B (Takeuchi et al., 1976 J. Antibiotics, 29, 93-96), porothramycin (Tsunakawa et al., 1988 J. Antibiotics, 41, 1366-1373), prothracarcin (Shimizu et al, 1982 J. Antibiotics, 29, 2492-2503; Langley and Thurston, 1987 J. Org. Chem., 52, 91-97), sibanomicin (DC-102) (Hara et al., 1988 J. Antibiotics, 41, 702-704; Itoh et al., 1988 J. Antibiotics, 41, 1281-1284), sibiromycin (Leber et al., 1988 J. Am. Chem. Soc., 110, 2992-2993) and tomamycin (Arima et al., 1972 J. Antibiotics, 25, 437-444).
PBDs are of the general structure: 
They differ in the number, type and position of substituents, in both their aromatic A rings and pyrrolo C rings, and in the degree of saturation of the C ring. There is either an imine (Nxe2x95x90C), a carbinolamine (NHxe2x80x94CH(OH)) or a carbinolamine methyl ether (NHxe2x80x94CH(OMe))at the N10-C11 position which is the electrophilic centre responsible for alkylating DNA. All of the known natural products have an (S)-configuration at the chiral C11a position which provides them with a right-handed twist when viewed from the C ring towards the A ring. This gives them the appropriate three-dimensional shape for isohelicity with the minor groove of B-form DNA, leading to a snug fit at the binding site (Kohn, 1975 in Antibiotics III. Springer-Verlag, New York, pp. 3-11; Hurley and Needham-VanDevanter, 1986 Acc. Chem. Res., 19, 230-237). Their ability to form an adduct in the minor groove, enables them to interfere with DNA processing, hence their use as antitumour agents.
A first aspect of the present invention relates to compounds of formula (I): 
wherein:
R2 and R3 are independently selected from: H, R, OH, OR, xe2x95x90O, xe2x95x90CH-R, xe2x95x90CH2, CH2xe2x80x94CO2R, CH2xe2x80x94CO2H, CH2xe2x80x94SO2R, Oxe2x80x94SO2R, CO2R, COR and CN, and there is optionally a double bond between C1 and C2 or C2 and C3;
R6, R7, R8, and R9, are independently selected from H, R, OH, OR, halo, nitro, amino, Me3Sn; or R7 and R8 together from a group xe2x80x94Oxe2x80x94(CH2)pxe2x80x94Oxe2x80x94, where p is 1 or 2;
R11, is either H or R;
Q is S, O or NH;
L is a linking group, or less preferably a single bond;
O is a solid support;
where R is a lower alkyl group having 1 to 10 carbon atoms, or an alkaryl group (i.e. an alkyl group with one or more aryl substituents) preferably of up to 12 carbon atoms, whereof the alkyl group optionally contains one or more carbon-carbon system, or an aryl group, preferably of up to 12 carbon atoms; and is optionally substituted by one or more halo, hydroxy, amino, or nitro groups, and optionally contains one or more hetero atoms, which may form part of, or be, a functional group; and
where one or more of R2, R3, R6, R7 and R8 may alternatively be independently Xxe2x80x94Yxe2x80x94Axe2x80x94, where X is selected from xe2x80x94COZxe2x80x2, NHZ, SH, or OH, where Z is either H or a nitrogen protecting group, Zxe2x80x2 is either OH or an acid protecting group, Y is a divalent group such that HY-R, and A is O, S, NH, or a single bond.
If R is an aryl group and contains a hetero atom, then R is a heterocyclic group. If R is an alkyl chain, and contains a hetero atom, the hetero atom may be located anywhere in the alkyl chain, e.g. xe2x80x94Oxe2x80x94C2H5, xe2x80x94CH2xe2x80x94Sxe2x80x94CH3, or may form part of, or be, a functional group, e.g. carbonyl, hydroxy, cyano, ester.
R and HY groups are preferably independently selected from a lower alkyl group having 1 to 10 carbon atoms, or an aralkyl group, preferably of up to 12 carbon atoms, or an aryl group, preferably of up to 12 carbon atoms, optionally substituted by one or more halo, hydroxy, amino, or nitro groups. It is more preferred that R and HY groups are independently selected from a lower alkyl group having 1 to 10 carbon atoms optionally substituted by one or more halo, hydroxy, amino, or nitro groups. It is particularly preferred that R or HY are unsubstituted straight or branched chain alkyl groups, having 1 to 10, preferably 1 to 6, and more preferably 1 to 4, carbon atoms, e.g. methyl, ethyl, propyl, butyl. R may be selected only from methyl and ethyl.
Alternatively, R6, R7, R8 and R9 may preferably be independently selected from R groups with the following structural characteristics:
(i) an optionally substituted phenyl group;
(ii) an optionally substituted ethenyl group;
(iii) an ethenyl group conjugated to an electron sink.
The term xe2x80x98electron sinkxe2x80x99 means a moiety-covalently attached to a compound which is capable of reducing electron density in other parts of the compound. Examples of electron sinks include cyano, carbonyl and ester groups.
The term xe2x80x98nitrogen protecting groupxe2x80x99 has the meaning usual in synthetic chemistry, particularly synthetic peptide chemistry. It means any group which may be covalently bound to the nitrogen atom of any grouping of the molecule, particularly of the amine grouping, and permits reactions to be carried out upon the molecule containing this protected grouping without its removal. Nevertheless, it is able to be removed from the nitrogen atom without affecting the remainder of the molecule. Suitable amine protecting groups for the present invention include Fmoc (9-fluorenylmethoxycarbonyl), Nvoc (6-nitroveratryloxycarbonyl), Teoc (2-trimethylsilylethyloxycarbonyl), Troc (2,2,2-trichloroethyloxycarbonyl), Boc (t-butyloxycarbonyl), CBZ (benzyloxycarbonyl), Alloc (allyloxycarbonyl) and Psec (2(-phenylsulphonyl)ethyloxycarbonyl). Other suitable groups are described in Protective Groups in Organic Synthesis, T Green and P Wuts, published by Wiley, 1991 which is incorporated herein by reference.
The term xe2x80x98acid protecting groupxe2x80x99 has the meaning usual in synthetic chemistry. It means any group which may be reacted with any carboxylic acid moiety of the molecule, and permits reactions to be carried out upon the molecule containing this protected grouping without its removal. Nevertheless, the carboxylic acid moiety is able to be regenerated without affecting the remainder of the molecule. Suitable acid protecting groups include esters, for example methyl ester, and xe2x80x94Oxe2x80x94CH2xe2x95x90CH2. Other suitable groups are described in Protective Groups in Organic Synthesis, T Green and P Wuts, published by Wiley, 1991.
It is preferred that in compounds of formula I, if one of R2, R3, R6, R7 and R8 is to be Xxe2x80x94Yxe2x80x94Axe2x80x94, then it is either R2 or R8 that is Xxe2x80x94Yxe2x80x94Axe2x80x94, and more preferably it is R8 that is Xxe2x80x94Yxe2x80x94Axe2x80x94.
In compounds of formula I, Q is preferably O, and R11 is preferably H, Me or ET, more preferably H or Me. Independently, R6 is preferably H or R, more preferably H or Me, R9 is preferably H, and R7 is preferably an alkoxy group, and more preferably methoxy or ethoxy. It is further preferred that R2 and R3 are H.
If there is a double bond in the pyrrolo C ring, it is preferably between C2 and C3.
A second aspect of the invention relates to compounds of formula I as defined in the first aspect of the invention except that one or more of R2, R3, R6, R7 and R8 are independently:
Hxe2x80x94(T)nxe2x80x94Xxe2x80x2xe2x80x94Yxe2x80x94Axe2x80x94
where:
Y and A are as defined in the first aspect of the invention;
Xxe2x80x2 is CO, NH, S or O,;
T is a combinatorial unit;
and n is a positive integer.
In compounds of formula I according to the second aspect, it is preferred that R2 and/or R8 are independently:
Hxe2x80x94(T)nxe2x80x94Xxe2x80x2xe2x80x94Yxe2x80x94Axe2x80x94
It is preferred that Xxe2x80x2 is either CO or NH. n may preferably be from 1 to 16, and more preferably from 3 to 14. It is also preferred that it is R8 which is Hxe2x80x94(T)nxe2x80x94Xxe2x80x2xe2x80x94Yxe2x80x94Axe2x80x94.
A third aspect of the present invention relates to compounds of formula II: 
preferably made from a compound of formula I as described in the first or second aspect of the invention by removing the compound of formula II from the solid support by cleaving the linking group L, where R2, R3, R6, R7, R8, and R9 are as defined in the first or second aspect of the invention.
A fourth aspect of the present invention is a method of making a compound according to the third aspect of the invention from a compound of formula I as described in the first or second aspect of the invention by removing the compound of formula II from the solid support by cleaving the linking group L.
A fifth aspect of the invention relates to a compound of formula II as described in the third aspect of the invention for use in a method of therapy. Conditions which may be treated include gene-based diseases, including neoplastic diseases and, for example Alzheimer""s disease, and bacterial, parasitic and viral infections.
In accordance with this aspect of the present invention, the compounds provided may be administered to individuals. Administration is preferably in a xe2x80x9ctherapeutically effective amountxe2x80x9d, this being sufficient to show benefit to a patient. Such benefit may be at least amelioration of at least one symptom. The actual amount administered, and rate and time-course of administration, will depend on the nature and severity of what is being treated. Prescription of treatment, e.g. decisions on dosages etc., is within the responsibility of general practitioners and other medical doctors.
A compound may be administered alone or in combination with other treatments, either simultaneously or sequentially dependent upon the condition to be treated.
Pharmaceutical compositions according to the present invention, and for use in accordance with the present invention, may comprise, in addition to the active ingredient, i.e. a compound of formula II, a pharmaceutically acceptable excipient, carrier, buffer, stabiliser or other materials well known to those skilled in the art. Such materials should be non-toxic and should not interfere with the efficacy of the active ingredient. The precise nature of the carrier or other material will depend on the route of administration, which may be oral., or by injection, e.g. cutaneous, subcutaneous or intravenous.
Pharmaceutical compositions for oral administration may be in tablet, capsule, powder or liquid form. A tablet may comprise a solid carrier or an adjuvant. Liquid pharmaceutical compositions generally comprise a liquid carrier such as water, petroleum, animal or vegetable oils, mineral oil or synthetic oil. Physiological saline solution, dextrose or other saccharide solutions, or glycols such as ethylene glycol, propylene glycol or polyethylene glycol may be included. Capsules may include a solid carrier such as gelatin.
For intravenous, cutaneous or subcutaneous injection, or injection at the site of affliction, the active ingredient will be in the form of a parenterally acceptable aqueous solution which is pyrogen-free and which has suitable pH, isotonicity and stability. Those of relevant skill in the art are well able to prepare suitable solutions using, for example, isotonic vehicles such as Sodium Chloride Injection, Ringer""s Injection or Lactated Ringer""s Injection. Preservatives, stabilisers, buffers, antioxidants and/or other additives may be included, as required.
A sixth aspect of the present invention relates to the use of a compound of formula II as described in the third aspect of the present invention in the preparation of a medicament for the treatment of a gene-based disease or a bacterial, parasitic or viral infection. The preparation of a medicament is described in relation to the fourth aspect of the invention.
In further aspects, the invention provides processes for preparing compounds according to the first and second aspects of the present invention.
The term xe2x80x98solid supportxe2x80x99 refers to a material having a rigid or semi-rigid surface which contains or can be derivatized to contain reactive functionality which can serve to covalently link a compound to the surface thereof. Such materials are well known in the art and include, by way of example, silicon dioxide supports containing reactive Si-OH groups, polyacrylamide supports, polystyrene supports, polyethyleneglycol supports, and the like. Such supports will preferably take the form of small beads, pins/crowns, laminar surfaces, pellets or disks. Other conventional forms may be used.
The linking groups preferred for the present application are ones which contain at least one covalent bond which can be readily broken by specific chemical reactions, or other changes (e.g. light or a pH change), thereby providing for liberation of compounds free from the solid support. The chemical reactions employed to break the covalent bond are selected so as to be specific for the desired bond breakage thereby preventing unintended reactions occurring elsewhere in the molecule. The linking group is selected relative to the synthesis of the compounds formed on the solid support so as to prevent premature cleavage of the compound or its precursors from the solid support as well as to avoid interference with any of the procedures employed during synthesis of the compound on the support.
Examples of linking groups are set out below (shown as available form), along with suggested cleavage method(s) for the linking group. These groups are commercially available or have been reported in the literature. After conversion to the appropriate chloroformate, for example by reaction with triphosgene in the presence of pyridine, they can be used to attach to anthranilic acids (for use in providing the protected A-rings of pyrrolobenzodiazepines) via carbamate linkages. Some resins, e.g. p-nitrophenyl carbonate Wang resin may couple to the anthranilic acids without need for intermediate transformation to the chloroformate. 
1. Holmes, C. P., Jones, D. G., xe2x80x9cReagents for Combinatorial Organic Synthesis: Development of a New O-Nitrobenzyl Photolabile Linker for Solid Phase Synthesisxe2x80x9d, J. Org. Chem., 60, 2318-2319 (1995).
2. Hauske, J. R., Dorff, P. A., xe2x80x9cSolid Phase CBZ Chloride Equivalent. A New Matrix Specific Linkerxe2x80x9d, Tetrahedron Letters, 36, 10, 1589-1592 (1995).
3. Kunz, H., Dombo, B., xe2x80x9cSolid Phase Synthesis of Peptide and Glycopeptides on Polymeric Supports with Allylic Anchor Groupsxe2x80x9d, Angew Chem Int Ed Engl, 5, 711 (1988).
4. Garcia-Echeverria, C., xe2x80x9cA Base Labile Handle for Solid Phase Organic Chemistryxe2x80x9d, Tetrahedron Letters, 38, 52, 8933-8934 (1997).
5. (a) Albericio, F., Giralt, E., Eritja, R., Tetrahedron Letters, 32, 1515 (1991).
(b) Albericio, F., Robles, J., Fernandez-Forner, Y., Palom, C., Celma, E., Pedroso, E., Giralt, E., Eritja, R., Peptides 1990, Proc 21st Eur. Pept. Symp., S134, (1991).
6. Mullen, D. G, Barany, G., xe2x80x9cA New Fluoridolyzable Anchoring Linkage for orthogonal Solid-Phase Peptide Synthesis; Design, Preparation, and Application of the N-(3 or 4)-[[4- (Hydroxymethyl)phenoxy]-tert-butylphenylsilyl]phenyl Pentanedioic Acid Monoamide (Pbs) Handlexe2x80x9d, J. Org. Chem., 53, 5240 (1988).
7. Dressman, D. A., et al., Tet. Letts., 37, 937 (1996). All these documents are incorporated herein by reference.
The term xe2x80x98combinatorial unitxe2x80x99 means any monomer unit which can be used to build a chain as shown in a compound of formula I as defined in the second aspect of the present invention, or a compound of formula II, when derived from a compound of formula I as defined in the second aspect of the present invention. The chain is usually attached to the PBD core by a joining group through the pro N10 position. Examples of molecules suitable for such chain building are found in Schreiber et al. (JACS, 120, 1998, pp.23-29), which is incorporated herein by reference. An important example of a unit is an amino acid residue. Chains may be synthesised by means of amine-protected amino acids. Fmoc protected amino-acids are available from a number of sources, such as Sigma and NovaBiochem. Both natural and unnatural amino acids can be used, e.g. D- and L-amino acids and heterocyclic amino acids. In particular, heterocyclic amino acids of the type found in the construction of netropsin and distamycin are of interest because of their DNA-recognition properties.
Amine units can be used to make up peptoids: see Soth, M. J. and Nowick, J. S. 1997, Unnatural oligomer libraries, Curr. Opin, Chem. Biol. 1, no. 1: 120-129; Zuckermann et al., 1994, Discovery of Nanomolecular Ligands for 7-Transmembrane G-Protein-Coupled Receptors from a Diverse N-(Substituted)glycine Peptoid Library, Journal of Medicinal Chemistry 37: 2678-85; Figliozzi, GMR et al., 1996, Synthesis of N-substituted Glycine Peptoid Libraries, Methods in Enzymology, 267: 437-47; Simon, R. J. et al., 1992, Peptoids: A Modular Approach to Drug Discovery, Proc. Natl. Acad. Sci. USA, 89:9367-71; which documents are incorporated herein by reference.
Other combinatorial units include PNAs: P E Nielsen, et al., Science, 1991, 254, 1497; M Egholm, et al., Nature, 1993, 365, 566; M Egholm et al., JACS, 1992, 114, 1895; S C Brown, et al., Science, 1994, 265, 777; 5. K Saha, et al., JOC, 1993, 58, 7827; oligoureas: K Burgess, et al., 1995, Solid Phase Synthesis of Unnatural Biopolymers Containing Repeating Urea Units. Agnew. Chem. Int. Ed. Engl 34, no. 8:907; K Burgess, et al., 1997, Solid Phase Synthesis of Oligoureas; Journal of the American Chemical Society 119: 1556-64; and oligocarbamates: E J Moran et al., 1995, Novel Biopolymers for Drug Discovery. Biopolymers (Peptide Science); John Wiley and Sons 37: 213-19; Cho C Y et al., 1993, An Unnatural Biopolymer. Science 261: 1303-5; Paikoff S F et al., 1996, The Solid Phase Synthesis of N-Alkylcarbamate Oligomers. Tetrahedron Letters 37, no. 32: 5653-56. All these documents are incorporated herein by reference.
A type of combinatorial unit of particular relevance to the present invention is one based on the pyrrolobenzodiazepine structures; these are of general formulae IIIa and IIIb: 
wherein R3, R6, R7 R9, A and Y are as defined in the first aspect of the invention, Axe2x80x2 and Yxe2x80x2 are independently selected from the possible groups for A and Y respectively. In order for such combinatorial units to be added to the combinatorial chain, they may be added in their protected form as shown in general formulae IIIc and IIId: 
where R3, R6, R7, R9, A, Y, Axe2x80x2 and Yxe2x80x2 are as defined above, Q and R11 are as defined in the first aspect of the invention, and R10 is a nitrogen protecting group. It is possible that the combinatorial units may remain in their protected form until the compound has been cleaved from the solid support, or until any other components of the compound have been deprotected.
The present invention relates to libraries, or collections, of compounds all of which are represented by a single one of the formulae I or II. The diversity of the compounds in a library may reflect the presence of compounds differing in the identities of one or more of R2, R3, R6, R7, R9, R11 and Q and/or in the identities of the combinatorial units T (when present). The number of members in the library depends on the number of variants, and the number of possibilities for each variant. For example, if it is the combinatorial units which are varied, and there are 3 combinatorial units, with 3 possibilities for each unit, the library will have 27 compounds. 4 combinatorial units and 5 possibilities for each unit gives a library of 625 compounds. If for instance there is a chain of 5 combinatorial units with 17 possibilities for each unit, the total number of members in the library would be 1.4 million. A library may therefore comprise more than 1 000, 5 000, 10 000, 100 000 or a million compounds, which may be arranged as described below.
In the case of free compounds (formula II), the individual compounds are preferably in discrete volumes of solvents, e.g. in tubes or wells. In the case of bound compounds (formula I) the individual compounds are preferably bound at discrete locations, e.g. on respective pins/crowns or beads. The library of compounds may be provided on a plate which is of a suitable size for the library, or may be on a number of plates of a standard size, e.g. 96 well plates. If the number of members of the library is large, it is preferable that each well on a plate contains a number of related compounds from the library, e.g. from 10 to 100. One possibility for this type of grouping of compounds is where only a subset of the combinatorial units, or substituents, are known and the remainder are randomised; this arrangement is useful in iterative screening processes(see below). The library may be presented in other forms that are well-known.
A further aspect of the present invention is a method of preparing a collection, or library of compounds as discussed above. If the diversity of the library is in the combinatorial units, then the library may be synthesised by the stepwise addition of protected combinatorial units to a PBD core, each step being interposed by a deprotection step. Such a method is exemplified later. If the diversity of the library is in the substituent groups, the library may be synthesised by carrying out the same synthetic methods on a variety of starting materials or key intermediates, which already possess the necessary substituent patterns.
The present invention also relates to a method of screening the compounds of formula II to discover biologically active compounds. The screening can be to assess the binding interaction with nucleic acids, e.g. DNA or RNA, or proteins, or to assess the affect of the compounds against protein-protein or nucleic acid-protein interactions, e.g. transcription factor DP-1 with E2F-1, or estrogen response element (ERE) with human estrogen receptor (a 66 kd protein which functions as a hormone-activated transcription factor, the sequence of which is published in the art and is generally available). The screening can be carried out by bringing the target macromolecules into contact with individual compounds or the arrays or libraries of individual compounds described above, and selecting those compounds, or wells with mixtures of compounds, which show the strongest effect.
This effect may simply be the cytotoxicity of the compounds in question against cells or the binding of the compounds to nucleic acids. In the case of protein-protein or nucleic acid-protein interactions, the effect may be the disruption of the interaction studied.
Protein-protein interactions can be measured in a number of ways, e.g. FRET (fluorescence resonance energy transfer) which involves labelling one of the proteins with a fluorescent donor moiety and the other with an acceptor which is capable of absorbing the emission from the donor; the fluorescence signal of the donor will be altered depending on the interaction between the two proteins. Another method of measuring protein-protein interactions is by enzymatic labelling, using, for example, horseradish peroxidase.
The screening process may undergo several iterations by selecting the most active compounds, or groups of compounds, tested in each iteration; this is particularly useful when testing arrays of wells which include mixtures of related compounds. Furthermore, if the wells contain compounds for which only a subset of the combinatorial units, or substituents, are known, but the rest are randomised, subsequent iterations can be carried out by synthesising compounds possessing the selected known (and successful) combinatorial unit, or substituent, pattern, but with further specified combinatorial units, or substituents, replacing the previously randomised combinatorial units, or substituents, adjacent the already known pattern; the remaining combinatorial units, or substituents, are randomised as in the previous iteration. This iterative method enables the identification of active members of large libraries without the need to isolate every member of the library.
A further feature of this aspect is formulation of a selected compound or selected compounds with pharmaceutically acceptable carriers or diluents.
A further aspect of the present invention relates to the use of compounds of formula II in target validation. Target validation is the disruption of an identified DNA sequence to ascertain the function of the sequence, and a compound of formula II can be used to selectively bind an identified sequence, and thus disrupt its function.
Compounds of formula II can also be used in functional genomics to ascertain the biological function of individual genes, by blocking this biological action. This is a further aspect of the invention.
A key step in a preferred route to compounds of formula I is a cyclisation procedure to produce the B-ring, involving generation of an aldehyde (or functional equivalent thereof) at what will be the 11-position, and attack thereon by the pro-10-nitrogen: 
The xe2x80x9cmasked aldehydexe2x80x9d xe2x80x94CPQ may be an acetal or thioacetal, which may be cyclic, in which case the cyclisation involves unmasking. Alternatively, it may be an alcohol xe2x80x94CHOH, in which case the reaction involves oxidation, e.g. by means of TPAP or DMSO (Swern oxidation).
The masked aldehyde compound can be produced by condensing a corresponding 2-substituted pyrrolidine with a 2-nitrobenzoic acid: 
The nitro group can then be reduced to xe2x80x94NH2 and reacted with a suitable linking group attached to a solid support, e.g. a chloroformate, which thereby links the structure to the solid support.
A process involving the oxidation-cyclization procedure is illustrated in scheme 1 (an alternative type of cyclisation will be described later with reference to scheme 2). 
If R11 is other than hydrogen, the compound of formula I, may be prepared by direct etherification of the alcohol Ia. Compounds with Qxe2x95x90S can be prepared by treatment of the corresponding alcohol Ia with R11SH, and a catalyst (usually a Lewis Acid such as Al2O3). For compounds where Qxe2x95x90NH, these can be prepared by reacting an amine, R11NH, e.g. C3H7NH with the corresponding alcohol Ia normally with a catalyst, such as a Lewis Acid.
Exposure of the alcohol B to tetrapropylammonium perruthenate (TPAP)/N-methylmorpholine -oxide (NMO) over A4 sieves results in oxidation accompanied by spontaneous B-ring closure to afford the desired product. The TPAP/NMO oxidation procedure is found to be particularly convenient for small scale reactions while the use of DMSO-based oxidation methods, particularly Swern oxidation, proves superior for larger scale work (e.g.  greater than 1 g).
The uncyclized alcohol B may be prepared by the reaction of the amino alcohol C, generally in solution, with the linking group L attached to a solid support D. The linking group is preferably terminated with a chloroformate or acid chloride functionality. This reaction is generally carried out in the presence of a base such as pyridine (preferably 2 equivalents) at a low temperature (e.g. at 0xc2x0 C.).
The key amino alcohol C may be prepared by reduction of the corresponding nitro compound E, by choosing a method which will leave the rest of the molecule intact. For example, treatment of E with tin (II) chloride in a suitable solvent, e.g. refluxing methanol, generally affords, after the removal of the tin salts, the desired product C in high yield.
Exposure of E to hydrazine/Raney nickel avoids the production of tin salts and may result in a higher yield of C, although this method is less compatible with the range of possible C and A-ring substituents. For instance, if there is C-ring unsaturation (either in the ring itself, or in R2 or R3), this technique may be unsuitable.
The nitro compound of formula E may be prepared by coupling the appropriate o-nitrobenzoyl chloride to a compound of formula F, e.g. in the presence of K2CO3 at xe2x88x9225xc2x0 C. under a N2 atmosphere. Compounds of formula F can be readily prepared, for example by olefination of the ketone derived from L-trans-hydroxy proline. The ketone intermediate can also be exploited by conversion to the enol triflate for use in palladium-mediated coupling reactions.
The o-nitrobenzoyl chloride is synthesised from the o-nitrobenzoic acid (or alkyl ester, after hydrolysis) of formula G, which itself is prepared from the vanillic acid (or alkyl ester) derivative H. Many of these are commercially available and some are disclosed in Althuis, T. H. and Hess, H. J., J. Medicinal Chem., 20(1), 146-266 (1977).

In scheme 1, the final or penultimate step was an oxidative cyclisation. An alternative route, using thioacetal coupling, is shown in scheme 2. Mercury-mediated unmasking causes cyclisation to the desired compound (Ia)
The thioacetal compound may be prepared as shown in scheme 2: the thioacetal protected C-ring [prepared via a literature method: Langley, D. R., and Thurston, D. E., J. Organic Chemistry, 52, 91-97 (1987)] is coupled to the o-nitrobenzoic acid (or alkyl ester) G using a literature procedure. The resulting nitro compound cannot be reduced by hydrogenation because of the thioacetal group, so the tin(II) chloride method is used to afford the amine. This is then N-protected, e.g., by reaction with a chloroformate or acid chloride, such as p-nitrobenzylchloroformate.
Acetal containing C-rings can be used as an alternative in this type of route with deprotection including other methods, including the use of Lewis Acid conditions (see example 3).
In the above synthesis schemes, the derivatisation of the A-ring is shown as being complete before the compounds are attached to the solid support. This is preferred if the substituents are groups such as alkoxy or nitro. On the other hand, substituent groups such as alkyl or alkenyl could be added to the A-ring after the coupling of the compound to the solid support. This may be achieved by R6, R7, R8, or R9 being easily replaceable groups, such as a halogen atom.
An alternative synthesis route (as in Examples 3 and 4xe2x80x94FIGS. 4 and 5) is to attach the component which will form the A ring to the solid support at the pro N10 position, before joining the component which will form the C ring.